MHC class II restricted antigen processing by Antigen Presenting Cells (APCs) is essential for the development of CD4+ T-cell responses. In this proposal we wish to investigate the involvement in this process of proteins which are resident in the intracellular MIIC class II compartments (MIICs) where the generation of MHC class II-peptide complexes occurs. The first, defined in the previous grant period, is a gamma interferon-inducible lysosomal thiolreductase (GILT) which is constitutively expressed in the MIICs of APCs. We hypothesize that GILT is involved in reducing the disulfide bonds within internalized protein antigens which must be broken to allow efficient antigen processing. The second is a subset of the tetraspan family of membrane proteins which resides within MIICs. The prototypical member, also identified in the previous funding period, is CD82. These proteins associate with MHC class II proteins, as well as the related proteins HLA-DM and HLA-DO, which regulate peptide loading within MIICs. We hypothesize that these proteins are involved in the maturation process which segregates class II molecules from resident proteins with the MIIC and therefore regulates the surface expression of MHC class II proteins. We propose to identify substrate proteins which interact with GILT, and to investigate the immunological and physiological functions of GILT using GILT-/- mice. We also propose to investigate the nature of tetraspan interactions with MHC class II molecules and other MIIC proteins, and to determine the role of the tetraspan proteins in MHC class II function at the biochemical level. These studies will unravel novel features of MHC class II-restricted antigen processing which may prove of clinical importance in immunological diseases, particularly autoimmunity.